The major objective for this laboratory is to develop technology capable of sequencing large genomes. We are pursuing a technology for sequencing individual DNA molecules using single-channel recording techniques that may significantly improve overall sequencing rates and read lengths. We propose that detectable changes in the conductance state of a channel protein will be caused by nucleotide bases in DNA, and that as DNA passes through or over the opening of the pore, these conductance states can be used to read the nucleotide sequence. Although this proposal only deals with very limited feasibility experiments, we want to consider reasonable limits for a refined device should feasibility be established. With computer data transfer rates on the order of Mbytes per second, a chip with 5000 operational pores reading at a rate of 200 bases/sec each and with associated data reduction circuit elements could read two genome equivalents from one human cell in two hours. This could allow far more accurate medical correlations than the present diagnostics that are based on a few select oligonucleotides. Even use of this technique for biopsy samples and microbial population sampling may be feasible. 1) We will monitor the conductance of single maltoporin (LamB) pores with high resolution single-channel recording techniques in a reconstituted system. 2) We will monitor the conductance of LamB pores during phage lambda injection of normal and modified DNA. 3) We will construct fusion proteins between LamB and T7 RNA polymerase such that both proteins retain their function, and then measure the conductance of individual pore-polymerase complexes with added template containing modified bases and the T7 RNA polymerase promoter. 4) We will analyze the effects of various modified nucleotides on pore conductance. We are after a reproducible correlation between specific modified nucleotides and LamB conductance states. 5) We will attempt to integrate promising modifications into a single system, so that all four nucleotides can be correlated with unique conductance states.